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Information on EC 1.13.11.20 - cysteine dioxygenase and Organism(s) Rattus norvegicus and UniProt Accession P21816
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1.13.11.20 cysteine dioxygenaseIUBMB Comments
Requires Fe2+ and NAD(P)H.
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Word Map
- 1.13.11.20
- taurine
-
sulfinic
-
non-heme
- hypotaurine
-
cysteinesulfinate
- cysteamine
- cystathionine
- 3-mercaptopropionate
-
cys-tyr
-
taut
-
adenosyltransferase
-
desulfhydration
-
2-his-1-carboxylate
-
gamma-lyase
-
cupins
- 2-aminoethanethiol
- medicine
The taxonomic range for the selected organisms is: Rattus norvegicus
The enzyme appears in selected viruses and cellular organisms
The enzyme appears in selected viruses and cellular organisms
Synonyms
cysteine dioxygenase, cysteine oxidase, cysteine dioxygenase type 1, bscdo, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
cysteine oxidase
-
-
-
-
oxygenase, cysteine di-
-
-
-
-
REACTION
REACTION DIAGRAM
COMMENTARY
ORGANISM
UNIPROT
LITERATURE
L-cysteine + O2 = 3-sulfinoalanine
catalytic mechanism of cysteine dioxygenase compared to the mechanism of sulfoxide synthase EgtB, EC 1.14.99.50, and its function of the active-site Tyr377 residue. In the sulfoxide syntase reaction, the conserved tyrosine residue reacts via proton-coupled electron transfer with the iron(III)-superoxo species and creates an iron(III)-hydroperoxo intermediate, thereby preventing the possible thiolate dioxygenation side reaction, detailed overview
L-cysteine + O2 = 3-sulfinoalanine
proposed catalytic cycle of CDO enzymes: the pentacoordinated resting state structure can convert to a hexacoordinated structure by binding of water. Upon binding of molecular oxygen the water molecule is displaced and an iron(III)-superoxo structure is formed. The terminal oxygen atom of the iron(III)-superoxo group attacks the sulfur of the cysteinate group to form a ring-structure, which can homogeneously split into a sulfoxide and iron(IV)-oxo species. The sulfoxide-bound complex rotates from sulfur-bound to oxygen-bound to the iron center. The latter transfers the oxygen atom from the iron(IV)-oxo group to substrate in a final step to form cysteine sulfinic acid product complex
L-cysteine + O2 = 3-sulfinoalanine
Upon binding substrate, the structure of the iron site is pertubed but is still consistent with six O/N donor, indicating that cysteine may bind to the iron center, but is not bound via the sulfur atom.
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
redox reaction
-
-
-
-
oxidation
-
-
-
-
reduction
-
-
-
-
PATHWAY SOURCE
PATHWAYS
BRENDA
-
-, -
SYSTEMATIC NAME
IUBMB Comments
L-cysteine:oxygen oxidoreductase
Requires Fe2+ and NAD(P)H.
CAS REGISTRY NUMBER
COMMENTARY
37256-59-0
-
SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
L-cysteine + O2
3-sulfinoalanine
-
-
-
?
L-cysteine + O2
3-sulfinoalanine
anaerobic CDO reaction conditions
-
-
?
L-cysteine + O2
3-sulfinoalanine
enzyme CDO-L-cysteine complex structure analysis and modeling, O2 binding structure, QM-MM simulations, detailed overview
-
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
-
-
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
highly specific for L-cysteine
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
highly specific for L-cysteine
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme of cysteine metabolism
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme of cysteine metabolism
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
liver enzyme responds to dietary protein contents, role in regulation of intracellular levels of methionine, cysteine and glutathione
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
enzyme expression in the brain may have several possible functions, like the prevention of free radical production by the autooxidation of cysteine and dopamine
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme in sulfate production, critical regulator of cellular cysteine concentration and availability of cysteine for anabolic processes
-
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme of cysteine catabolism, supplies substrate for taurine biosynthesis
-
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme of taurine biosynthesis, provides substrate for transamination, regulation of intracellular cysteine level
-
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
regulation of intracellular cysteine concentration
-
-
?
additional information
?
-
formation of an active CDO:cysteine substrate complex
-
-
?
additional information
?
-
homocysteine and D-Cys are no substrates
-
-
?
additional information
?
-
-
glutathione, dithiothreitol and cystine do not serve as substrates
-
-
?
additional information
?
-
-
L-cystine, D-cysteine, DL-homocysteine and cysteamine do not serve as substrates
-
-
?
additional information
?
-
-
L-cystine, D-cysteine, carboxymethyl-L-cysteine, carboxyethyl-L-cysteine, S-methyl-L-cysteine, N-acetyl-L-cysteine, DL-homocysteine and cysteamine do not serve as substrates
-
-
?
additional information
?
-
-
cysteamine does not serve as substrate, D-cysteine does not serve as substrate
-
-
?
additional information
?
-
-
cysteine catabolism in mammals is dependent upon cysteine dioxygenase. System for regulation of cellular cysteine levels. Evidence of abnormal or deficient CDO activity has been reported in individuals with a variety of autoimmune and neurodegenerative diseases, including rheumatoid arthritis, Parkinsons disease, Alzheimers disease, and motor neuron diseases
-
-
?
additional information
?
-
-
cysteine dioxygenase cannot catalyze the oxidation of selenocysteine. In the Cys-bound complexes, the change of the oxidation state for the Fe center is II to III to II, while the Fe center in the Sec-bound complexes remains in the II oxidation state throughout. The competition for donation of electron density with the iron ion determines the valence change and the reaction ability
-
-
?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
L-cysteine + O2
3-sulfinoalanine
-
-
-
?
additional information
?
-
-
cysteine catabolism in mammals is dependent upon cysteine dioxygenase. System for regulation of cellular cysteine levels. Evidence of abnormal or deficient CDO activity has been reported in individuals with a variety of autoimmune and neurodegenerative diseases, including rheumatoid arthritis, Parkinsons disease, Alzheimers disease, and motor neuron diseases
-
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme of cysteine metabolism
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme of cysteine metabolism
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
liver enzyme responds to dietary protein contents, role in regulation of intracellular levels of methionine, cysteine and glutathione
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
enzyme expression in the brain may have several possible functions, like the prevention of free radical production by the autooxidation of cysteine and dopamine
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme in sulfate production, critical regulator of cellular cysteine concentration and availability of cysteine for anabolic processes
-
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme of cysteine catabolism, supplies substrate for taurine biosynthesis
-
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
key enzyme of taurine biosynthesis, provides substrate for transamination, regulation of intracellular cysteine level
-
-
?
L-cysteine + O2
3-sulfino-L-alanine
-
regulation of intracellular cysteine concentration
-
-
?
COFACTOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
additional information
enzyme uses an amino acid cofactor in the active site consisting of two cross-linked residues, cysteine 93 and tyrosine 157. Formation of the Cys-Tyr cofactor requires a transition metal cofactor Fe2+ and O2. Biogenesis of the cofactor is also strictly dependent upon the presence of substrate. In the absence of the Cys-Tyr cofactor, the enzyme possesses appreciable catalytic activity. At physiologically relevant cysteine concentrations, cofactor formation increases catalytic efficiency 10-fold
-
additional information
-
NAD+ does not appear to be a cofactor, the role of NAD+ in the stimulation of the enzyme is unclear
-
additional information
-
no requirement of NAD, NADP, NADH or NADPH for the enzymic activity of protein-B. The enzymic activity of protein-B alone is extremely low, protein-A alone does not exhibit catalytic activity, however a significant activity is observed in the presence of both fractions, requirement of protein-A for the catalytic activity of protein-B
-
additional information
-
NADPH or NADH do not act as cosubstrates, most of added NADPH disappears from the incubation mixture during the first 10 min, while cysteinesulfinic acid remains linear for up to 30 min, the role of NADHP or NADH may be one of stabilization, as allosteric activators or other type of activating agents
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Fe2+
Ni2+
the optimized structure of the dioxygen-bound active site features an S=2 quintet ground state, with a Ni(III)-superoxo species
Fe
Fe2+
Iron
-
loosely bound to protein, only 10% of purified protein contains iron
Fe2+
required for activity, an iron-oxygen intermediate is formed during the catalytic cycle of cysteine dioxygenase, Moessbauer spectroscopy analysis resuting in three different species: CDO:FeII, CDO:FeII:cysteine site I, and CDO:FeII:cysteine site II, dissociation constants, absorptions spectra, and structure analysis, detailed overview
Fe2+
required, enzyme-bound. The iron ion is coordinated by H86, H88, H140 and a water molecule, and can further accept a coordinating L-cysteine molecule
Fe
-
Fe2+ required, 0.8-0.9 gatom of iron per 22500 g of purified protein
Fe2+
-
additon of 0.2 or 0.3 mM Fe2+ yields near-optimal activity. Activity of fully purified enzyme in the absence of added Fe2+ is below 1% of that measured in the presence of 0.3 mM ferrous sulfate.
Fe2+
-
required for catalysis, in the Cys-bound complexes, the change of the oxidation state for the Fe center is II to III to II. Binding involves coordination with His86, His88, and His140
Fe2+
-
strongly bound high-spin iron(II) coordinates cysteine and homocysteine in cysteine dioxygenase, overview. CDO tightly binds iron(II) at a 3His active site. Although the cross-link between C93 and Y157 is close to the active site, it does not appear to affect iron binding. Stopped-flow measurements and Mössbauer spectral analysis, overview
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
2-amino-ethanethiol
1 x the Km for cysteine = 8.7% inhibition, 10 x the Km for cysteine = 30% inhibition
2-sulfanyl-ethanol
1 x the Km for cysteine = 5.9% inhibition, 10 x the Km for cysteine = 13% inhibition
3-sulfanyl-propionic acid
1 x the Km for cysteine = 4.9% inhibition, 10 x the Km for cysteine = 26% inhibition
azide
competitive inhibitor, binding structure, overview. Azide does not bind in the enzyme crystal as a superoxide mimic
alpha-ketoglutarate
-
alpha-ketoglutarate inhibits cysteine dioxygenase with 50% inhibition at 6.8 mM
aspartic acid
-
aspartic acid decreases enzyme activity to 50% at a concentration of 1.5 mM. Replacing the sulfydryl by the uncharged hydroxyl group of serine does not affect enzyme activity.
CdCl2
-
cells transfected with wild-type enzyme show an enhanced sensitivity to CdCl2 that is limited to cells cultured in medium with cysteine levels of 0.1 and 0.3mM
D-cysteinesulfinate
-
34.4% inhibition in hepatocytes from rats fed a low casein diet, 71.8% inhibition in hepatocytes from rats fed a moderate casein diet, 64.4% inhibition in hepatocytes from rats fed a high casein diet
EGTA
-
with protein-A: 100% inhibition at 0.1 mM, without protein-A: 95% inhibition at 0.1 mM
L-cysteine
-
concentrations of cysteine of 2 mM and above are inhibitory in assays of purified cysteine dioxygenase
Mercaptopropionic acid
-
mercaptopropionic acid at a concentration of 1.2 mM inhibits cysteine dioxygenase activity by 50%
Neocuproine
-
with protein-A: 18% inhibition at 0.1 mM, without protein-A: slight activation at 0.1 mM
S-carboxymethylcysteine
-
S-carboxymethylcysteine exhibits 50% inhibition at a concentration of 2.3 mM
2,2'-dipyridyl
-
with protein-A: 100% inhibition at 0.1 mM, without protein-A: 58% inhibition at 0.1 mM
2,2'-dipyridyl
-
non preincubated protein-B: 76% inhibition at 1 mM, with cysteine preincubated protein-B: no inhibition
8-hydroxyquinoline
-
with protein-A: 100% inhibition at 0.1 mM, without protein-A: 99% inhibition at 0.1 mM
Bathocuproine sulfonate
-
80% inhibition at 0.01 mM, 83% inhibition at 0.1 mM
Bathocuproine sulfonate
-
with protein-A: 5% inhibition at 0.1 mM, without protein-A: 38% inhibition at 0.1 mM
bathophenanthroline sulfonate
-
68% inhibition at 0.01 mM, 100% inhibition at 0.1 mM
bathophenanthroline sulfonate
-
with protein-A: 88% inhibition, without protein-A: 100% inhibition
DL-propargylglycine
-
45.5% inhibition in hepatocytes from rats fed a high casein diet
DL-propargylglycine
-
reduces the enzyme activity in methionine-supplemented medium to the basal level, does not reduce the enzyme activity in cysteine-supplemented medium, no effect in hepatocytes cultured in basal medium
EDTA
-
non preincubated protein-B: 91% inhibition at 1 mM, with cysteine preincubated protein-B: 51% inhibition at 1 mM
EDTA
-
with protein-A: 97% inhibition at 0.1 mM, with protein-A: 99% inhibition at 0.1 mM
Fe2+
-
inhibits the enzyme activity of both preactivated and non-preactivated protein-B
o-phenanthroline
-
non-preincubated protein-B: 91% inhibition at 1 mM, with cysteine preincubated protein-B: 67% inhibition at 1 mM
o-phenanthroline
-
without protein-A: 96% inhibition at 0.1 mM, with protein A: 100% inhibition at 0.1 mM
additional information
-
iodoacetamide has no effect on the enzyme activity
-
additional information
-
S-methylcysteine does not inhibit cysteine dioxygenase
-
additional information
-
inhibitors of the 26S proteasome (e.g., proteasome inhibitor 1 and lactacystin) block CDO degradation in cysteine-deficient cells but had little or no effect on CDO concentration in hepatocytes cultured with excess cysteine
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
cysteine
-
protein expression of recombinant wild-type enzyme in HepG2/C3A cells increases by 160% when extracellular cysteine levels are increased from 0 to 1 mM cysteine
cysteamine
-
at a reaction time of 45 min, CDO activity increases almost 20times in the presence of 5.8 mM cysteamine
L-cysteine
-
activates the purified enzyme under anaerobic conditions
L-cysteine
-
acts as an initial signal for regulation of the enzyme, upregulates enzyme activity
additional information
the enzyme activity is in part modulated by the formation of a Cys93-Tyr157 crosslink that increases its catalytic efficiency over 10fold, mechanism, overview. The crosslink enhances activity by positioning the Tyr157 hydroxyl to enable proper Cys binding, proper oxygen binding, and optimal chemistry
-
additional information
-
cysteine mediates the up-regulation of enzyme in cultured hepatocytes
-
additional information
-
with pure cysteine dioxygenase no effect of addition of NAD+ to the cysteine diosygenase is found.
-
additional information
-
effect of dietary protein or cystine on CDO expression and enzyme activity in rat liver and adipose tissue
-
additional information
-
covalent post-translational modification between the residues C93 and Y157, in close proximity to the active site, enhances the enzyme's activity. The presence of ferrous iron and oxygen is a prerequisite for C93-Y157 crosslink formation. Both the enzymatic rate of cysteine oxidation and the amount of cross-linking between C93 and Y157 increased significantly upon exposure of CDO to air/oxygen and substrate cysteine in the presence of iron in a hitherto unreported two-phase process. The non-crosslinked form has negligible enzymatic activity
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.002
L-cysteine
20% cross-linked wild-type enzyme, pH 9.1, temperature not specified in the publication
0.003
L-cysteine
not cross-linked wild-type enzyme, pH 7.7, temperature not specified in the publication
0.005
L-cysteine
60% cross-linked wild-type enzyme, pH 7.7, temperature not specified in the publication
1.5
L-cysteine
measurement of cysteine sulfinic acid production is done by high-performance liquid chromatography
4.1
L-cysteine
-
purified recombinant enzyme, in 10 mM MES (pH 6.1) and 20 mMNaCl, at 37°C
additional information
additional information
stopped-flow kinetics of wild-type and mutant enzymes
-
additional information
additional information
steady-state Michaelis-Menten kinetics
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.012
L-cysteine
-
assay standart conditions are: the enzyme is incubated at 37°C in the presence of 62.5 mM Mes buffer (pH 6.1), 0.3 mM ferrous sulfate, 0.0125 mM bathocuproine disulfonate and 1.2 mM cysteine
0.12
L-cysteine
20% cross-linked wild-type enzyme, pH 9.1, temperature not specified in the publication
0.25
L-cysteine
60% cross-linked wild-type enzyme, pH 7.7, temperature not specified in the publication
0.33
L-cysteine
not cross-linked wild-type enzyme, pH 7.7, temperature not specified in the publication
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.056
L-cysteine
20% cross-linked wild-type enzyme, pH 9.1, temperature not specified in the publication
0.056
L-cysteine
60% cross-linked wild-type enzyme, pH 7.7, temperature not specified in the publication
0.095
L-cysteine
not cross-linked wild-type enzyme, pH 7.7, temperature not specified in the publication
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
0.000068
-
in isolated hepatocytes from rats fed the diet containing a low casein level
0.00016
-
in hepatocytes isolated from rats fed diets containing 100 g casein/kg without sulfur amino acid supplementation
0.00017
-
in hepatocytes isolated from rats fed diets containing 100 g casein/kg with 2.4 g L-cystine per kg supplementation
0.0003
-
in isolated hepatocytes from rats fed the diet containing a moderate casein level
0.00046
-
in hepatocytes isolated from rats fed diets containing 100 g casein/kg with 3 g L-methionine per kg supplementation
0.00047
-
in isolated hepatocytes from rats fed the diet containing a high casein level
0.0012
-
in hepatocytes isolated from rats fed diets containing 100 g casein/kg with 8 g L-cystine per kg supplementation
0.00202
-
in hepatocytes isolated from rats fed diets containing 100 g casein/kg with 10 g L-methionine per kg supplementation
additional information
additional information
-
enzyme activity measured in liver homogenate, supernatant and activated supernatant with and without addition of Fe2+, NAD+ and NH2OH. Comparison of enzyme activity in liver from Wistar vs Sprague-Dawley rats in the soluble and particulate fractions of liver homogenized in sucrose vs Mes buffer. Comparison of enzyme activity in hepatocytes isolated from rats fed diets with either 100 or 300 g casein per kg
additional information
-
With Lineweaver-Burk plot Vmax is estimated to be 1.870 micromol cysteinesulfinate/min/mg
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
6.1
pH optimum is determined by using 2-morpholinoethanesulfonic acid or Tris buffers at a final concentration of 62.5 mM
6.8 - 9.5
-
for the anaerobic activation of the purified enzyme by L-cysteine
7.5
additional information
pH-dependence of wild-type and mutant enzyme kinetics, detailed overview
pH RANGE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
37
38 - 40
-
for the anaerobic activation of the purified enzyme by L-cysteine
pI VALUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
Uniprot
native and selenomethionine-substituted cysteine dioxygnease
Uniprot
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
-
CDO is found specifically in the mucus-secreting goblet cells
-
intense staining for CDO in ductal cells of pregnant rats but not in other mammary epithelial cells or in ductal cells of non-pregnant rats
additional information
-
distribution is found to be centrilobular and does not alter when the enzyme is induced with cysteine or methionine
-
neuron, including the pyramidal cells of the hippocampus and the Purkinje cells of the cerebellum, the regional localization varies, with high levels of expression in the hippocampus, the dentate gyrus, the outer cortices of the brain, and the substantia nigra
-
primary cell culture, high enzyme activity in cysteine rich medium
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
evolution
structure and catalytic mechanism comparisons of nonheme iron enzymes cysteine dioxygenase with sulfoxide synthase EgtB, EC 1.14.99.50, quantum mechanics/molecular mechanics calculations, overview
metabolism
the O2 activation mechanism suggests the binding of O2 to the metal ion followed by the attack of the distal oxygen atom on the cysteine sulfur. An alternative mechanism entails the attack of the cysteine sulfur on the proximal oxygen atom of the dioxygen moiety to form a persulfenate intermediate without any redox exchange between the metal ion and the O2 ligand. The O2 activation mechanism with a Ni-substituted active site follows the same pattern as native CDOs albeit with much higher energy barriers for the formation of the intermediates. The immediate cleavage of the persulfenate S-O bond in the alternative mechanism suggests that cysteine persulfenate might not be a true intermediate
physiological function
additional information
physiological function
cysteine dioxygenase (CDO) helps regulate Cys levels through converting Cys to Cys sulfinic acid. The enzyme activity is in part modulated by the formation of a Cys93-Tyr157 crosslink that increases its catalytic efficiency over 10fold, mechanism, overview. The crosslink enhances activity by positioning the Tyr157 hydroxyl to enable proper Cys binding, proper oxygen binding, and optimal chemistry
physiological function
in the ferrous form, CDO catalyzes irreversibly the conversion of cysteine to cysteine sulfinate, incorporating both atoms of dioxygen into the product. Cysteine sulfinate is an intermediate of several pathways related to pyruvate and sulfurated metabolites, such as sulfate, taurine, and hypotaurine. The CDO performance increases in response to high cysteine levels,either through the formation of a C93-Y157 crosslink, or a diminished degradation by the ubiquitin-proteasome system
additional information
persulfenate and persulfide binding in the active site of cysteine dioxygenase, overview
additional information
modeling the pH-dependence of cysteine dioxygenation
additional information
structure modeling of cysteine dioxygenase in complex with L-cysteine dianion
additional information
the CDO enzyme active site structure comprises residues Tyr58, Arg60, His86, His88, Cys93, His140, and Tyr157, structure model
additional information
the structure of CDO, which is highly conserved across multiple species, is built on a small alpha-helical domain containing three alpha-helices at the N-terminus, followed by 13 beta-strands. These are subdivided into a main beta-sandwich domain and two beta-hairpins at the C terminus. The entire beta-sandwich is composed of seven anti-parallel beta-strands on the lower side and six anti-parallel beta-strands on the upper side. The active site comprises an iron ion, which is located in the central portion of the cupin beta-sandwhich, and is connected to the bulk solvent through a solvent-filled channel
additional information
wild-type and mutant active site structures, overview
additional information
-
covalent post-translational modification between the residues C93 and Y157, in close proximity to the active site, enhances the enzyme's activity. The presence of ferrous iron and oxygen is a prerequisite for C93-Y157 crosslink formation. Both the enzymatic rate of cysteine oxidation and the amount of cross-linking between C93 and Y157 increased significantly upon exposure of CDO to air/oxygen and substrate cysteine in the presence of iron in a hitherto unreported two-phase process. The non-crosslinked form has negligible enzymatic activity
additional information
-
structures of iron-containing CDO model complexes, modeling, overview
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). CDO1_RAT
200
0
23026
Swiss-Prot
other Location (Reliability: 2)
PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
22500
23000
23030
23500
23830
-
calculated from deduced amino acid sequence, His-tag fusion protein
68000
-
detection in liver whole homogenate, immunoabsorption of anti-enzyme antibodies
additional information
-
no 68000 Da species detected as reported in different publications
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
monomer
additional information
-
liver enzyme is composed of 2 distinct proteins: 1. protein-B, tightly bound iron as prosthetic group, 2. protein A, modifier or activating protein
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
additional information
Cys93-Sgamma is covalently bound to Tyr157-Cepsilon2 forming a cysteinyl-tyronsine linkage.
additional information
residues C93-Y157 crosslink as a result of a posttranslational modification
additional information
the enzyme activity is in part modulated by the formation of a Cys93-Tyr157 crosslink that increases its catalytic efficiency over 10fold, mechanism, overview. The crosslink enhances activity by positioning the Tyr157 hydroxyl to enable proper Cys binding, proper oxygen binding, and optimal chemistry
additional information
-
two posttranslational modifications adjacent to the catalytic iron center: a thioether cross-link between Cys93 and Tyr157 and extra electron density at Cys164 which is variously explained as cystine or cysteine sulfinic acid
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
crystallization in sitting drops at 25°C using a reservoir of 0.1-0.25 M ammonium acetate, 0.1 M tri-sodium citrate, pH 5.6, with 22-26% (w/v) polyethylene glycol 4000. The co-crystals with 5 mM are grown using a reservoir of 0.15 M ammonium sulfate, 0.2 M sodium cacodylate, pH 6.5, with 26% (w/v) polyethylene glycol 8000. The crystal sturcture, solved by SAD phasing using selenomethionine-substituted protein, yields a final refined model with r = 18.0 and Rfree = 20.8 at 1.5-A resolution. Data from a co-crystallization experiment with 5 mM cysteine shows structural changes in the binding pocket, they are determined to 1.5 A resolution (final r = 19.8 and Rfree = 22.4).
generation of 21 CDO crystal structures at resolutions between 1.25 and 1.65 A. Of these, 16 are of C93A or Y157F CDO mutants either alone or bound to L-Cys, D-Cys, or the inhibitor homocysteine, the other five are of wild-type CDO bound to homocysteine, azide, or thiosulfate. Cys-soaked wild-type CDO crystals are analysed at pH 6.2 and pH 8.0, detailed overview
purified enzyme, hanging drop vapour diffusion method, mixing of 0.0015 ml of 7 mg/mL CDO mutant C93G in 10 mM sodium phosphate and 20 mM NaCl, pH 7.5, and 0.0006 ml of crushed wild-type CDO crystal seeds in their own growth solution consisting of 25% w/v PEG 1500, 13 mM succinate, 44 mM sodium phosphate, and 44 mM glycine, pH 5.2, with 0.0015 ml of reservoir buffer containing 26% w/v PEG 4000, 200 mM ammonium acetate, and 100 mM sodium citrate, pH 6.3, equilibration against reservoir solution, X-ray diffraction structure determination and analysis at 1.82 A resolution
purified recombinant enzyme complexed with cysteine persulfide or 3-mercaptopropionic acid persulfide, hanging drop vapor diffusion method, mixing of 0.0015 ml of 8 mg/ml protein with 0.0015 ml of reservoir solution containing 24-34% w/v PEG 4000, 100-250 mM ammonium acetate, 100 mM sodium citrate, pH 5.6, 0-4 mM dithionite, and 40 mM ligand, final pH is 6.1-6.2, 24°C, one week, X-ray diffraction structure determination and analysis at 1.63-2.05 A resolution
structures of C93E and cross-linked and non-cross-linked wild-type CDO, to 1.91, 2.49, and 2.30 A, respectively. Mutant C93E has similar overall structural properties compared to cross-linked CDO, but the iron is coordinated by a 3-His/1-Glu geometry. The hydroxyl group of Tyr157 shifts in both non-cross-linked and C93E CDO, and this displacement prevents the residue from participating in substrate stabilization
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
C93A
site-directed mutagenesis, mutation of the active site residue, no crosslink formation resulting in reduced activity compared to the wild-type enzyme. The mutant variant structure has a new chloride ion coordinating the active site iron. Cys binding is also different from wild-type CDO, and no Cys-persulfenate forms in the C93A active site at pH 6.2 or pH 8.0
C93E
mutation to the corresponding residue of cupin to reestablish the common 3-His/1-Glu metal ligand of the cupin superfamily. Mutant shows dioxygen consumption, which, is not coupled with L-cysteine oxidation. Substrate analogues (D-cysteine, cysteamine, and 3-mercaptopropionate) show variable coordinations to the iron center, but are not viable substrates for the variant
C93G
Y157F
site-directed mutagenesis, mutation of the active site residue, no crosslink formation resulting in reduced activity compared to the wild-type enzyme. The mutant variant structure has a new chloride ion coordinating the active site iron. Cys binding is also different from wild-type CDO, and no Cys-persulfenate forms in the Y157F active site at pH 6.2 or pH 8.0
C93S
-
mutant can not be converted to the mature form due to the loss of the cysteine residue involved in thioether crosslink formation
H86A
R60A
-
mutant forms with low activity, which has a markedly decreased affinity for cysteine, probably due to the loss of the hydrogen bonding partner for the carboxylate of the substrate, forms the crosslink more slowly
additional information
C93G
site-directed mutagenesis, structure comparison to the wild-type enzyme
H86A
-
expression of either the wild-type or a catalytically inactivated mutant H86A in HepG2/C3A cells which do not express endogenous cysteine dioxygenase protein and culture in different concentrations of extracellular cysteine. Wild-type enzyme, but not mutant H86A, is capable of reducing intracellular cysteine levels in cells incubated in physiologically relevant concentrations of cysteine. Wild-type enzyme also decreases the glutathione pool and potentiates the toxicity of CdCl2
H86A
-
inactive mutant, leading to loss of the His that serves as a metal ligand in the active site does not form any crosslink
additional information
a selenomethionine-substituted cysteine dioxygenase is produced
additional information
C93A and Y157F unliganded active site structures comparison
additional information
-
recombinant cysteine dioxygenase protein with a thioredoxin sequence and 6 x His tagged, after purification the 6x His tag and the thioredoxin sequence are removed using factor Xa.
OXIDATION STABILITY
ORGANISM
UNIPROT
LITERATURE
rapid and irreversible inactivation under aerobic conditions, inactivation can be prevented by a distinct cytoplasmic protein, i.e. protein A
-
439542, 439546, 439548
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
all buffers are supplemented with 5 mM dithiothreitol to prevent oxidation of the selenomethionine
recombinant Strep-tagged wild-type and mutant enzyme by affinity chromatography to over 95% purity
recombinant thioredoxin-His6-tagged enzyme by affinity chromatography
1 ml HisTrap HP column, metal ion (nickel) affinity chromatography, gel filtration, MonoQ 4.6/100 PE ion exchange column. The thioredoxin/6x His cysteine dioxygenase fusion protein separates into two apparent isoforms that elute as two distinct peaks, one that elutes at 50 mM imidazole and one that elutes at 100 mM imidazole during metal ion affinity chromatograohy. The protein in the second peak has a specific activity, that is 50-60% less than that of the protein in the first peak. In contrast to peak 1 peak 2 elutes as two pronounced peaks (A and B) from the MonoQ column. The cysteine dioxygenase in peak A has no detectabel activity. In the the standard cysteine dioxygenase purification procedure, only the form from peak 1 is retained and further purified.
-
recombinnat enzyme from Escherichia coli strain BL21(DE3) with cleavage of the Thx-His6 tag
-
using acetone fractionation, column chromatography on DEAE-cellulose, Sephadex G-100, hydroxylapatite, DEAE-Sephadex A-25 and Sephadex G-75
-
using acetone precipitation, first chromatography on DEAE-cellulose column, second chromatography on DEAE-cellulose column, chromatography on Sephadex G-100 column, hydroxyapatite column, DEAE-Sephadex A-25 column and Sephadex G-75 column
-
using acid treatment, ammonium sulfate fractionation and column chromatography with DEAE-cellulose. The purified enzyme is composed of two distinct proteins, it appears that one of them is a catalytic protein named protein-B having tightly bound iron as a prosthetic group, while the other is either a modifier or activating protein named protein-A. Protein-B is found to exist in both an active and an inactive form
-
using heat treatment, ammonium sulfate fractionation and column chromatography on DEAE-cellulose, Sephadex G-200 and Sephadex G-100
-
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene cdo, expression of Thx-His6 ´-tagged enzyme in Escherichia coli strain BL21(DE3)
-
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Hosokawa, Y.; Matsumoto, A.; Oka, J.; Itakura, H.; Yamaguchi, K.
Isolation and characterization of a cDNA for rat liver cysteine dioxygenase
Biochem. Biophys. Res. Commun.
168
473-478
1990
Rattus norvegicus
Yamaguchi, K.; Hosokawa, Y.
Cysteine dioxygenase
Methods Enzymol.
143
395-403
1987
Rattus norvegicus
Lombardini, J.B.; Singer, T.P.; Boyer, P.D.
Cystein oxygenase. II. Studies on the mechanism of the reaction with 18oxygen
J. Biol. Chem.
244
1172-1175
1969
Rattus norvegicus
Kohashi, N.; Yamaguchi, K.; Hosokawa, Y.; Kori, Y.; Fujii, O.; Ueda, I.
Dietary control of cysteine dioxygenase in rat liver
J. Biochem.
84
159-168
1978
Rattus norvegicus
Sakakibara, S.; Yamaguchi, K.; Hosokawa, Y.; Kohashi, N.; Ueda, I.; Sakamoto, Y.
Purification and some properties of rat liver cysteine oxidase (cysteine dioxygenase)
Biochim. Biophys. Acta
422
273-279
1976
Rattus norvegicus
Sakakibara, S.; Yamaguchi, K.; Ueda, I.
Two components of cysteine oxidase in rat liver
Biochem. Biophys. Res. Commun.
52
1093-1099
1973
Rattus norvegicus
Yamaguchi, K.; Hosokawa, Y.; Kohashi, N.; Kori, Y.; Sakakibara, S.; Ueda, I.
Rat liver cysteine dioxygenase (cysteine oxidase). Further purification, characterization, and analysis of the activation and inactivation
J. Biochem.
83
479-491
1978
Rattus norvegicus
Bagley, P.J.; Stipanuk, M.H.
The activities of rat hepatic cysteine dioxygenase and cysteinesulfinate decarboxylase are regulated in a reciprocal manner in response to dietary casein level
J. Nutr.
124
2410-2421
1994
Rattus norvegicus
McCann, K.P.; Akbari, M.T.; Williams, A.C.; Ramsden, D.B.
Human cysteine dioxygenase type I: primary structure derived from base sequencing of cDNA
Biochim. Biophys. Acta
1209
107-110
1994
Homo sapiens, Rattus norvegicus
Bagley, P.J.; Stipanuk, M.H.
Rats fed a low protein diet supplemented with sulfur amino acids have increased cysteine dioxygenase activity and increased taurine production in hepatocytes
J. Nutr.
125
933-940
1995
Rattus norvegicus
Bagley, P.J.; Hirschberger, L.L.; Stipanuk, M.H.
Evaluation and modification of an assay procedure for cysteine dioxygenase activity: high-performance liquid chromatography method for measurement of cysteine sulfinate and demonstration of physiological relevance of cysteine dioxygenase activity in cysteine catabolism
Anal. Biochem.
227
40-48
1995
Rattus norvegicus
Parsons, R.B.; Ramsden, D.B.; Waring, R.H.; Barber, P.C.; Williams, A.C.
Hepatic localization of rat cysteine dioxygenase
J. Hepatol.
29
595-602
1998
Rattus norvegicus
Kwon, Y.H.; Stipanuk, M.H.
Cysteine regulates expression of cysteine dioxygenase and gamma-glutamylcysteine synthetase in cultured rat hepatocytes
Am. J. Physiol. Endocrinol. Metab.
280
E804-815
2001
Rattus norvegicus
Parsons, R.B.; Waring, R.H.; Williams, A.C.; Ramsden, D.B.
Cysteine dioxygenase: regional localization of protein and mRNA in rat brain
J. Neurosci. Res.
65
78-84
2001
Rattus norvegicus
Stipanuk, M.H.; Hirschberger, L.L.; Londono, M.P.; Cresenzi, C.L.; Yu, A.F.
The ubiquitin-proteasome system is responsible for cysteine-responsive regulation of cysteine dioxygenase concentration in liver
Am. J. Physiol.
286
E439-448
2004
Rattus norvegicus
Stipanuk, M.H.; Londono, M.; Hirschberger, L.L.; Hickey, C.; Thiel, D.J.; Wang, L.
Evidence for expression of a single distinct form of mammalian cysteine dioxygenase
Amino Acids
26
99-106
2004
Rattus norvegicus
Chai, S.C.; Jerkins, A.A.; Banik, J.J.; Shalev, I.; Pinkham, J.L.; Uden, P.C.; Maroney, M.J.
Heterologous expression, purification, and characterization of recombinant rat cysteine dioxygenase
J. Biol. Chem.
280
9865-9869
2005
Rattus norvegicus
Cresenzi, C.L.; Lee, J.I.; Stipanuk, M.H.
Cysteine is the metabolic signal responsible for dietary regulation of hepatic cysteine dioxygenase and glutamate cysteine ligase in intact rats
J. Nutr.
133
2697-2702
2003
Rattus norvegicus
Dominy, J.E.; Simmons, C.R.; Karplus, P.A.; Gehring, A.M.; Stipanuk, M.H.
Identification and characterization of bacterial cysteine dioxygenases: a new route of cysteine degradation for eubacteria
J. Bacteriol.
188
5561-5569
2006
Bacillus cereus (Q81CX4), Bacillus cereus, Bacillus cereus DSM 31 (Q81CX4), Bacillus subtilis (O32085), Bacillus subtilis, no activity in Nostoc sp., Rattus norvegicus (P21816), Streptomyces coelicolor (O50490), Streptomyces coelicolor (Q9KZL0), Streptomyces coelicolor A3(2) SCO3035 (Q9KZL0), Streptomyces coelicolor A3(2) SCO5772 (O50490)
Chai, S.C.; Bruyere, J.R.; Maroney, M.J.
Probes of the catalytic site of cysteine dioxygenase
J. Biol. Chem.
281
15774-15779
2006
Rattus norvegicus
Simmons, C.R.; Liu, Q.; Huang, Q.; Hao, Q.; Begley, T.P.; Karplus, P.A.; Stipanuk, M.H.
Crystal structure of mammalian cysteine dioxygenase. A novel mononuclear iron center for cysteine thiol oxidation
J. Biol. Chem.
281
18723-18733
2006
Rattus norvegicus (P21816)
Simmons, C.R.; Hirschberger, L.L.; Machi, M.S.; Stipanuk, M.H.
Expression, purification, and kinetic characterization of recombinant rat cysteine dioxygenase, a non-heme metalloenzyme necessary for regulation of cellular cysteine levels
Protein Expr. Purif.
47
74-81
2006
Rattus norvegicus
Dominy, J.E.; Hwang, J.; Stipanuk, M.H.
Overexpression of cysteine dioxygenase reduces intracellular cysteine and glutathione pools in HepG2/C3A cells
Am. J. Physiol. Endocrinol. Metab.
293
E62-E69
2007
Rattus norvegicus
Dominy, J.E.; Hwang, J.; Guo, S.; Hirschberger, L.L.; Zhang, S.; Stipanuk, M.H.
Synthesis of amino acid cofactor in cysteine dioxygenase is regulated by substrate and represents a novel post-translational regulation of activity
J. Biol. Chem.
283
12188-12201
2008
Rattus norvegicus (P21816)
Ueki, I.; Stipanuk, M.H.
3T3-L1 adipocytes and rat adipose tissue have a high capacity for taurine synthesis by the cysteine dioxygenase/cysteinesulfinate decarboxylase and cysteamine dioxygenase pathways
J. Nutr.
139
207-214
2009
Rattus norvegicus
Stipanuk, M.H.; Ueki, I.; Dominy, J.E.; Simmons, C.R.; Hirschberger, L.L.
Cysteine dioxygenase: a robust system for regulation of cellular cysteine levels
Amino Acids
37
55-63
2009
Rattus norvegicus
Kleffmann, T.; Jongkees, S.A.; Fairweather, G.; Wilbanks, S.M.; Jameson, G.N.
Mass-spectrometric characterization of two posttranslational modifications of cysteine dioxygenase
J. Biol. Inorg. Chem.
14
913-921
2009
Rattus norvegicus
Siakkou, E.; Wilbanks, S.M.; Jameson, G.N.
Simplified cysteine dioxygenase activity assay allows simultaneous quantitation of both substrate and product
Anal. Biochem.
405
127-131
2010
Rattus norvegicus
Tchesnokov, E.P.; Wilbanks, S.M.; Jameson, G.N.L.
A strongly bound high-spin iron(II) coordinates cysteine and homocysteine in cysteine dioxygenase
Biochemistry
51
257-264
2012
Rattus norvegicus
Souness, R.J.; Kleffmann, T.; Tchesnokov, E.P.; Wilbanks, S.M.; Jameson, G.B.; Jameson, G.N.
Mechanistic implications of persulfenate and persulfide binding in the active site of cysteine dioxygenase
Biochemistry
52
7606-7617
2013
Rattus norvegicus (P21816)
Siakkou, E.; Rutledge, M.T.; Wilbanks, S.M.; Jameson, G.N.L.
Correlating crosslink formation with enzymatic activity in cysteine dioxygenase
Biochim. Biophys. Acta
1814
2003-2009
2011
Rattus norvegicus
Che, X.; Gao, J.; Liu, Y.; Liu, C.
Metal vs. chalcogen competition in the catalytic mechanism of cysteine dioxygenase
J. Inorg. Biochem.
122
1-7
2013
Rattus norvegicus
Davies, C.G.; Fellner, M.; Tchesnokov, E.P.; Wilbanks, S.M.; Jameson, G.N.
The Cys-Tyr cross-link of cysteine dioxygenase changes the optimal pH of the reaction without a structural change
Biochemistry
53
7961-7968
2014
Rattus norvegicus (P21816)
Pietra, F.
On the dynamical behavior of the cysteine dioxygenase-L-cysteine complex in the presence of free dioxygen and L-cysteine
Chem. Biodivers.
14
e1700290
2017
Rattus norvegicus (P21816)
Tchesnokov, E.P.; Faponle, A.S.; Davies, C.G.; Quesne, M.G.; Turner, R.; Fellner, M.; Souness, R.J.; Wilbanks, S.M.; de Visser, S.P.; Jameson, G.N.
An iron-oxygen intermediate formed during the catalytic cycle of cysteine dioxygenase
Chem. Commun. (Camb.)
52
8814-8817
2016
Rattus norvegicus (P21816)
Faponle, A.S.; Seebeck, F.P.; de Visser, S.P.
Sulfoxide synthase versus cysteine dioxygenase reactivity in a nonheme iron enzyme
J. Am. Chem. Soc.
139
9259-9270
2017
Rattus norvegicus (P21816)
Driggers, C.M.; Kean, K.M.; Hirschberger, L.L.; Cooley, R.B.; Stipanuk, M.H.; Karplus, P.A.
Structure-based insights into the role of the Cys-Tyr crosslink and inhibitor recognition by mammalian cysteine dioxygenase
J. Mol. Biol.
428
3999-4012
2016
Rattus norvegicus (P21816)
Forbes, D.L.; Meneely, K.M.; Chilton, A.S.; Lamb, A.L.; Ellis, H.R.
The 3-His metal coordination site promotes the coupling of oxygen activation to cysteine oxidation in cysteine dioxygenase
Biochemistry
59
2022-2031
2020
Rattus norvegicus (P21816)
Attia, A.; Silaghi-Dumitrescu, R.
Nickel-substituted iron-dependent cysteine dioxygenase Implications for the dioxygenation activity of nickel model compounds
Int. J. Quantum Chem.
118
e25564
2018
Rattus norvegicus (P21816)
-
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